This invention relates generally to a novel assay method. More particularly, it relates to an assay method for the detection of direct bilirubin, also called conjugated bilirubin, in serum and other body fluids.
While it is typical in the field of diagnostic analysis to utilize the same compound assayed for as the standard in the clinical determination, e.g. glucose in assays for specimen glucose, it is not a practical solution in the case of direct bilirubin. This material is not only difficult ot obtain but it is relatively unstable.
The prior art has suggested the use of N-1-naphthyl ethylenediamine for the standardization of direct bilirubin assays. It has been found that while N-1-naphthyl ethylenediamine is suitable as a standard for the calibration of direct bilirubin assays in a single-assay system, it is not suitable for clinical analyzers which simultaneously analyze a single specimen for more than one component, particularly where ion-selective electrodes are used in the multi-assay system.
For instance, ion-selective electrodes are used for both sodium and potassium determinations. When N-1-naphthyl ethylenediamine is used for standardizing direct bilirubin in a multi-component system such as in the Technicon SMAC.sup.1 analyzer which requires compatibility with the other nineteen channels of this analyzer, it was found to be unsuitable because values for potassium were falsely elevated when potassium is determined by the normally-used valinomycin ion-selective electrode. As a result, there is a variable interference of N-1-naphthyl ethylenediamine (NEDD) in assays of potassium which varies from 0.1 - 1.5 meg./1 potassium for the desired level of 50 mg NEDD/1. FNT .sup.1 This is a Registered Trademark of Technicon Instruments Corporation.
A similar interference problem would be found for liquid membrane electrodes as well as in PVC membranes used in the ion-selective electrodes for assays of potassium.
It is therefore one object of this invention to provide a material available for standardization of assays for direct bilirubin in serum, suitable for inclusion in a multi-parameter serum without interference with the simultaneous assay of other components in the specimen, especially potassium.